Radiolabeled chelating compounds are useful both as medical diagnostic and therapeutic agents. For example, radiolabeled ethylenediamine tetraacetic acid (EDTA), and diethylenetriaminepentaacetate (DTPA) have been reported to be useful in evaluating renal functions, Klingensmith et al., J. Nucl. Med. 23:377 (1982). Similarly, Kasina et al., J. Med. Chem. 29:1933 (1986) report promising renal pharmaceuticals that are technetium chelates of N.sub.2 S.sub.2 diamido dimercaptides. Many other radiolabeled diagnostic chelates have been reported and include: tartrate and orthophosphate, Molinski et al., U.S. Pat. No. 3,987,1571 propylene amine oxime, Troutner et al., U.S. Pat. No. 4,615,876; polyhydroxycarboxylic acids, Adler et al., U.S. Pat. No. 4,027,005; organotrisubstituted trivalent phosphorus compounds, Dean et al., U.S. Pat. No. 4,582,700; bis-thiosemicarbazone, Vedee et al., U.S. Pat. No. 4,564,472; gentisyl alcohol in combination with phosphonates, Fawzi, U.S. Pat. No. 4,232,000; mercaptoacetylglycylglycylglycine (MAG.sub.3) Fritzberg et al., J. Nucl. Med. 27:111-116 (1986); mercaptocarboxylic acids, Winchell et al., U.S. Pat. No. 4,233,285; thiosaccharides, Kubiatowicz et al., U.S. Pat. No. 4,208,398; homocysteine and homocysteinamide derivatives, Byrne et al., U.S. Pat. No. 4,571,430; metallothionein, Tolman, European application Apr. 10, 1984 0 137 457 AZ; isonitrile, Jones et al., U.S. Pat. No. 4,452,774; and imidodiphosphonate, Subramanian et al., U.S. Pat. No. 3,974,268.
One class of such compounds is the bifunctional chelating compounds, which have a functional group capable of binding a metal and a functional group reactive with a carrier molecule. Compounds of this type are being actively investigated since they are capable of stably linking radionuclides to target-specific biological molecules such as proteins, antibodies, and antibody fragments.
Diagnostic imaging of specific target tissue in vivo with a radiometal-chelate-antibody conjugate was reported by Khaw et al., Science 209:295 (1980). Similarly, the therapeutic use of radiometal-chelate-antibody conjugates to treat cellular disorders is disclosed by Gansow et al., U.S. Pat. No. 4,454,106.
The procedure employed to insert a radiometal into a chelating compound depends on the chemistry of the radiometal and the chemical structure of the chelating compound. A variety of radiometals can be incorporated into both simple and bifunctional chelating compounds. The particular radiometal selected depends on the intended application and availability, as well as other factors.
Generally, radiometals intended for use as therapeutic agents are alpha, beta, or Auger electron emitters, such .sup.109 Pd, .sup.111 Ag, .sup.119 Sb, .sup.198 Au, .sup.199 Au, .sup.67 Cu, .sup.105 Rh, .sup.186 Re, .sup.188 Re, and .sup.212 Bi. Radiometals intended for use as diagnostic agents are usually positron or gamma photon emitters. For example, in positron emission tomography .sup.43 Sc, .sup.44 Sc, .sup.52 Fe, .sup.55 Co, and .sup.68 Ga can be employed, while for gamma camera imaging .sup.203 Pb, .sup.97 Ru, .sup.197 Hg, .sup.67 Ga, .sup.201 Tl, .sup.99m Tc, .sup.113m In, and .sup.111 In are usually selected.
Many of the radiometals described above are available in oxidation states unsuitable for chelation without prior treatment. .sup.99m Tc, for example, is available as pertechnetate (TcO.sub.4) and must be reduced to a lower oxidation state before chelation can occur. This is usually accomplished by the addition of a reducing agent, such as Sn.sup.+2 or dithionite at alkaline pH to the pertechnetate chelator mixture.
Transfer of the radiometal to the ultimate chelator is often facilitated by employing a labile or weak chelating agent (WCA) in the reaction mixture, Fritzberg et al. (1986). In the case of .sup.99m Tc, for example, an initial complex may be formed with a WCA such as gluconate. The .sup.99m Tc-gluconate complex forms quickly, thereby minimizing reoxidation of the .sup.99m Tc. Heating the initial Tc-WCA complex in the presence of a strong chelating agent (SCA) results in transfer of .sup.99m Tc to the strong chelating agent in improved yields, compared to carrying out the reduction of pertechnetate in the presence of the strong chelator alone.
Pollack et al. British J. Hematology, 34:231 (1976) describe the kinetic nature of the problem of transferring metals between strong chelating agents. These authors demonstrate a significantly enhanced transfer rate when a weak chelating agent, such as nitrilotriacetate, is employed.
The need to enhance the transfer kinetics of a metal to a strong chelator is particularly important when the chelator is attached to a protein. For example, Childs et al., J. Nucl. Med. 26:293-299 (1985) describe the rather harsh conditions, i.e. pH 4, necessary to achieve adequate binding of a radiometal to the antibody-bound chelator. Exposure to high temperatures or extremes of pH may denature or otherwise damage the protein to which the chelating compound is attached. Examples of weak chelating agents that have been used to facilitate transfer of metals to proteins or strong chelating agents attached thereto include the polyhydroxycarboxylates, glucoheptonate, Burchiel et al., J. Nucl. Med. 27:896 (1986) and tartrate, Kasina et al., Proc. Intl. Radio. Chem. Symp. 269-71 (1986). Strong chelating agents that have been conjugated to target specific proteins include: DTPA, Childs et al. (1985); EDTA, Wieder et al., U.S. Pat. No. 4,352,751 (1982); metallothionein, Tolman, European Patent Application 0137457 (1985); bis-thiosemicarbozones, Arano et al., Int. J. Nucl. Med. Bio. 12:425 (1986), U.S. Pat. No. 4,287,362; and diamido dimercaptide (N.sub.2 S.sub.2) Fritzberg et. al. (1986).
Even when a weak chelating agent is used to facilitate incorporation of a radiometal into a strong chelating compound, the kinetics are not always sufficient unless somewhat harsh conditions are employed. It is known, for example, that transfer of technetium from a Tc-tartrate complex to an antibody-N.sub.2 S.sub.2 conjugate is slow and requires heating to 50.degree. C. or more for an hour to effect acceptable radiometal transfer. See European Patent Application Publication No. 188,256. Heating to temperatures above 37.degree. C. often leads to aggregation of proteins such as antibodies, as well as nonspecific labeling of the antibody itself.
Accordingly, a need exists for a chelating compound that can rapidly form stable chelates with radiometals at physiological temperatures or below. Radiolabeling of bifunctional chelators suitable for conjugation to target-specific biological molecules should be possible under conditions that preserve biological activity.